Copper-catalyzed Azide-alkyne Cycloaddition Research Articles

MOFs Modulate Copper Trafficking in Tumor Cells for Bioorthogonal Therapy.

In situ drug synthesis using the copper-catalyzed azide-alkyne cycloaddition (CuAAC) reaction has attracted considerable attention in tumor therapy because of its satisfactory effectiveness and reduced side-effects. However, the exogenous addition of copper catalysts can cause cytotoxicity and has hampered biomedical applications in vivo. Here, we design and synthesize a metal-organic framework (MOF) to mimic copper chaperone, which can selectively modulate copper trafficking for bioorthogonal synthesis with no need of exogenous addition of copper catalysts. Like copper chaperones, the prepared ZIF-8 copper chaperone mimics specifically bind copper ions through the formation of coordination bonds. Moreover, the copper is unloaded under the acidic environment due to the dissipation of the coordination interactions between metal ions and ligands. In this way, the cancer cell-targeted copper chaperone mimics can selectively transport copper ions into cells. Regulation of intracellular copper trafficking may inspire constructing bioorthogonal catalysis system with reduced metal cytotoxicity in live cells.

On the mechanochemical synthesis of C-scorpionates with an oxime moiety and their application in the copper-catalyzed azide–alkyne cycloaddition (CuAAC) reaction

Integrating the principles of green chemistry to develop sustainable processes.

Dual-ligand PROTACS mediate superior target protein degradation in vitro and therapeutic efficacy in vivo.

Proteolysis targeting chimeras (PROTACs) are revolutionizing the drug development landscape due to their unique ability to selectively degrade disease-associated proteins. Conventional PROTACs are bivalent entities that induce ubiquitination and subsequent proteolysis of a chosen protein of interest (POI) by forming a ternary complex with an E3 ligase. We hypothesized that dual-ligand PROTACs, featuring two copies each of a POI ligand and an E3 ligase ligand, would facilitate the formation of high-avidity, long-lived ternary complexes inside cells, thereby increasing POI degradation potency. To this end, we developed a convergent synthesis route, using l-aspartic acid as a building block for homodimer synthesis, followed by copper-catalyzed azide-alkyne cycloaddition (CuAAC) to conjugate both dimers through a flexible linker. Dual-ligand PROTACs achieved up to a tenfold increase in degradation efficiency and a hundredfold increase in cytotoxicity in vitro across various cancer cell lines compared to their single-ligand counterparts. Furthermore, dual-ligand PROTACs sustain prolonged protein degradation, up to 60 hours after pulsing and washout. In vivo, in a mouse tumor model, the superior therapeutic activity of dual ligand PROTACs was observed.

Hydroquinone-treated Cu3(BTC)2: a mixed-valence Cu(I/II) MOF catalyst for efficient cycloadditions.

We present mixed-valence Cu(I)1Cu(II)2(BTC)2 [henceforth Cu(I/II)-HKUST-1], post-synthetically prepared via the hydroquinone (H2Q) treatment of Cu(II)3(BTC)2 (also referred to as HKUST-1) and its subsequent catalytic activity. This Cu(I/II)-HKUST-1 exhibits exceptional structural integrity and superior catalytic performance in the copper-catalyzed azide-alkyne cycloaddition (CuAAC) reaction between phenylacetylene and benzyl azide. These findings highlight the potential of mixed-valence Cu-based MOFs as effective and sustainable heterogeneous catalysts for organic transformations, paving the way for future advancements in MOF-based catalysis.

Iterative click reactions using trivalent platforms for sequential molecular assembly.

A facile synthesis of multi(triazole)s by iterative click reactions is disclosed. Good functional group tolerance of sequential click assembly by sulfur-fluoride exchange (SuFEx), copper-catalyzed azide-alkyne cycloaddition (CuAAC), and thia-Michael reaction realizes the iterative click reactions. Diverse multi(triazole)-type mid-molecules can be synthesized easily from readily available modules through good chemoselective reactions without functional group transformation steps.

Synthesis of substituted triazole-pyrazole hybrids using triazenylpyrazole precursors.

A synthesis route to access triazole-pyrazole hybrids via triazenylpyrazoles was developed. Contrary to existing methods, this route allows the facile N-functionalization of the pyrazole before the attachment of the triazole unit via a copper-catalyzed azide-alkyne cycloaddition. The developed methodology was used to synthesize a library of over fifty new multi-substituted pyrazole-triazole hybrids. We also demonstrate a one-pot strategy that renders the isolation of potentially hazardous azides obsolete. In addition, the compatibility of the method with solid-phase synthesis is shown exemplarily.

Real-time Visualization of Transcribed mRNA via Click Chemistry in a Liposomal Space.

We present a CuAAC (Copper-Catalyzed Azide-Alkyne Cycloaddition) reaction protocol designed for the visualization of mRNA. To achieve this, we synthesized stable mRNA molecules incorporating the modified nucleoside analog, EU, a crucial element for fluorophore attachment. Leveraging this modified mRNA, we successfully executed the CuAAC reaction, wherein the pro-fluorophore, coumarin, was conjugated to EU on the mRNA through our meticulously designed CuAAC process. This innovative approach resulted in the emission of fluorescence, enabling both precise quantification and visual observation of mRNA. Furthermore, we demonstrated the feasibility of concurrent mRNA synthesis and visualization by seamlessly integrating the CuAAC reaction mix into the mRNA transcription process. Additionally, our novel methodology opens avenues for prospective real-time monitoring of mRNA transcription within artificial cells. These advancements hold significant promise for expanding our comprehension of fundamental cellular processes and finding applications across diverse biological contexts in the future.

Cyclic Peptide Conjugate Vaccines and Physically Mixed Cyclic Peptide Vaccines for Subcutaneous Immunization.

Immune stimulants (adjuvants) enhance immune system recognition to provide an effective and individualized immune response when delivered with an antigen. Synthetic cyclic deca-peptides, co-administered with a toll-like receptor targeting lipopeptide, have shown self-adjuvant properties, dramatically boosting the immune response in a murine model as a subunit peptide-based vaccine containing group A Streptococcus peptide antigens.Here, we designed a novel peptide and lipid adjuvant system for the delivery of group A Streptococcus peptide antigen and a T helper peptide epitope. Following linear peptide synthesis on 2-chlorotrityl chloride resin, the linear peptide was cleaved and head-to-tail cyclized in solution. The selective arrangement of amino acids in the deca-peptide allowed for selective conjugation of lipids and/or peptide antigens following cyclisation. Using both solution-phase peptide chemistry and copper-catalyzed azide-alkyne cycloaddition reaction were covalently (and selectively) ligated lipid and/or peptide antigens onto the cyclic deca-peptide core. Subcutaneous administration of the vaccine design to mice resulted in the generation of a large number of serum immunoglobulin (Ig) G antibodies.

Coumarin-azasugar-benzyl conjugates as non-neurotoxic dual inhibitors of butyrylcholinesterase and cancer cell growth.

We have applied the copper-catalyzed azide-alkyne cycloaddition (CuAAC) reaction to prepare a library of ten coumarin-azasugar-benzyl conjugates and two phthalimide-azasugar-benzyl conjugates with potential anti-Alzheimer and anti-cancer properties. The compounds were evaluated as cholinesterase inhibitors, demonstrating a general preference, of up to 676-fold, for the inhibition of butyrylcholinesterase (BuChE) over acetylcholinesterase (AChE). Nine of the compounds behaved as stronger BuChE inhibitors than galantamine, one of the few drugs in clinical use against Alzheimer's disease. The most potent BuChE inhibitor (IC50 = 74 nM) was found to exhibit dual activities, as it also showed high activity (GI50 = 5.6 ± 1.1 μM) for inhibiting the growth of WiDr (colon cancer cells). In vitro studies on this dual-activity compound on Cerebellar Granule Neurons (CGNs) demonstrated that it displays no neurotoxicity.

Novel isatin-triazole based thiosemicarbazones as potential anticancer agents: synthesis, DFT and molecular docking studies.

Thiosemicarbazones of isatin have been found to exhibit versatile bioactivities. In this study, two distinct types of isatin-triazole hybrids 3a and 3b were accessed via copper-catalyzed azide-alkyne cycloaddition reaction (CuAAC), together with their mono and bis-thiosemicarbazone derivatives 4a-h and 5a-h. In addition to the characterization by physical, spectral and analytical data, a DFT study was carried out to obtain the optimized geometries of all thiosemicarbazones. The global reactivity values showed that among the synthesized derivatives, 4c, 4g and 5c having nitro substituents are the most soft compounds, with compound 5c having the highest electronegativity and electrophilicity index values among the synthesized series, thus possessing strong binding ability with biomolecules. Molecular docking studies were performed to explore the inhibitory ability of the selected compounds against the active sites of the anticancer protein of phosphoinositide 3-kinase (PI3K). Among the synthesized derivatives, 4-nitro substituted bisthiosemicarbazone 5c showed the highest binding energy of -10.3 kcal mol-1. These findings demonstrated that compound 5c could be used as a favored anticancer scaffold via the mechanism of inhibition against the PI3K signaling pathways.

Highly efficient functionalization of hydroxypropyl cellulose with glucuronic acid through click chemistry under microwave irradiation as potential biomaterial with therapeutic properties

A straightforward and effective method to chemically conjugate hydroxypropyl cellulose (HPC) to GlcA is described via microwave assisted copper-catalyzed azide-alkyne cycloaddition reaction (CuAAC) between the azide functionalized HPC and the alkynyl functionalized GlcA, reducing reaction times from 24 h to 2 h. Two types of HPC azide derivatives, including an ester (ES) or a carbamate (CB) functional group, were synthesized and characterized by FT-IR and NMR spectroscopy that revealed the efficiency in the incorporation of the azide moieties in the cellulose backbone. These compounds were then conjugated with the alkynyl GlcA by CuAAC, affording the desired glycopolymers, HPA-ES-GlcA and HPA-CB-GlcA in excellent yields. The whole process involved rapid and effective reactions as well as easy purification techniques, using mostly non-toxic solvents, contributing to the sustainability of the synthesis These glycopolymers were characterized by FT-IR, NMR, TGA, DTA and SEM. Furthermore, the cytotoxicity, anti-inflammatory and antibacterial properties of the new glycopolymers were tested and it was found that these new HPC-GlcA conjugates presented low cytotoxicity, good anti-inflammatory and antibacterial properties. Overall, our results showed that HPC-GlcA polymers are suitable as potential GAG mimics for biomedical applications, therefore further investigations will be carried out.

Copper-Catalyzed Azide-Alkyne Cycloaddition-Oriented Multifunctional Bio-Orthogonal Linker BPPA: Design, Synthesis and Evaluation.

The multifunctional linker molecules are crucial for the bio-orthogonal reaction for proteomic target profiling. Herein, we wish to present a novel type of biotin-based tetra-functional bio-orthogonal linkers 3a-3h named BPPA which, possessing a unique photolabile phenacyl ester motif, were readily prepared in 85-90% yields by a simple and green one-step protocol from commercially available and inexpensive reagents of biotin acids and 4'-ethynyl/azido 2-bromoacetophenones. The typical click reaction of BPPA linkers 3a and 3e via copper-catalyzed azide-alkyne cycloaddition (CuAAC) took place easily, resulting in the corresponding BPPA-triazole adducts 4a and 4b in nearly quantitative yields. A further cleavability evaluation of 4a and 4b demonstrated that the expected C-O bond detachment could be accomplished efficiently and rapidly by UV irradiation or by ammonia hydrolysis, respectively, resulting in the residual (hydroxyl)acetylphenyl triazole fragment supposed to be attached to proteins during biological manipulations. The BPPA linkers, with dual clickable options of either the terminal azide or alkyne clickable group, exhibit high potentials for various CuAAC-oriented bio-orthogonal reactions.

Click processes orthogonal to CuAAC and SuFEx forge selectively modifiable fluorescent linkers.

The appeal of catalytic click chemistry is largely due to the copper-catalysed azide-alkyne cycloaddition (CuAAC) process, which is orthogonal to the more recently introduced sulfur-fluoride exchange (SuFEx). However, the triazole rings generated by CuAAC are not readily modifiable, and SuFEx connectors cannot be selectively functionalized, attributes that would be attractive in a click process. Here we introduce bisphosphine-copper-catalysed phenoxydiazaborinine formation (CuPDF), a link-and-in situ modify strategy for merging a nitrile, an allene, a diborane and a hydrazine. We also present copper- and palladium-catalysed quinoline formation (Cu/PdQNF), which is applicable in aqueous media, involving an aniline as the modifier. CuPDF and Cu/PdQNF are easy to perform and deliver robust, alterable and tunable fluorescent hubs. CuPDF and Cu/PdQNF are orthogonal to SuFEx and CuAAC, despite the latter and CuPDF also being catalysed by an organocopper species. These advantages were applied to protecting group-free syntheses of sequence-defined branched oligomers, a chemoselectively amendable polymer, three drug conjugates and a two-drug conjugate.

Thymidine Kinase-Independent Click Chemistry DNADetect Probes for DNA Proliferation Assessment in Malaria Parasites.

Metabolic chemical probes are small-molecule reagents that utilize naturally occurring biosynthetic enzymes for in situ incorporation into biomolecules of interest. These reagents can be used to label, detect, and track important biological processes within living cells including protein synthesis, protein glycosylation, and nucleic acid proliferation. A limitation of current chemical probes, which have largely focused on mammalian cells, is that they often cannot be applied to other organisms due to metabolic differences. For example, the thymidine derivative 5-ethynyl-2'-deoxyuridine (EdU) is a gold standard metabolic chemical probe for assessing DNA proliferation in mammalian cells; however, it is unsuitable for the study of malaria parasites due to Plasmodium species lacking the thymidine kinase enzyme that is essential for metabolism of EdU. Herein, we report the design and synthesis of new thymidine-based probes that sidestep the requirement for a thymidine kinase enzyme in Plasmodium. Two of these DNADetect probes exhibit robust labeling of replicating asexual intraerythrocytic Plasmodium falciparum parasites, as determined by flow cytometry and fluorescence microscopy using copper-catalyzed azide-alkyne cycloaddition to a fluorescent azide. The DNADetect chemical probes are synthetically accessible and thus can be made widely available to researchers as tools to further understand the biology of different Plasmodium species, including laboratory lines and clinical isolates.

Influence of polymer amphiphilicity on the cyclization efficiency of poly(2-oxazoline)s

The steadily increasing interest in cyclic polymers has led to a plethora of proposed applications, particularly as biomaterials. However, the large-scale synthesis of cyclic polymers remains one of the greatest challenges in the field. One reason for this is the poorly understood role of the polymer structure on the cyclization efficiency, leading to synthesis protocols that must be optimized on a case-by-case basis without consideration for the chemical nature of the polymer. Recently, we found through a combination of laboratory experiments and molecular dynamics simulations that the polymer backbone flexibility has a major influence on the cyclization efficiency of poly(cyclic imino ether)s. Concurrently, we identified that a more flexible polymer backbone was also better solvated in the given organic solvent. To deconvolute the importance of the polymer backbone flexibility and solvation, in this study a variety of α-alkyne, ω-azide terminated (co)polymers of 2-ethyl-2-oxazoline and 2–n-butyl-2-oxazoline with different amphiphilicities, but the same polymer backbone, have been synthesized and subjected to copper-catalysed azide-alkyne cycloaddition (CuAAC) reactions in water. In agreement with our previous studies, it was observed that an increase in polymer chain hydrophobicity, and hence a decrease in solvation, leads to a significant decrease in cyclization efficiency. To further understand the effect of different CuAAC reaction conditions on the reaction outcome, different polymer concentrations and reaction temperatures were screened. This study highlights the importance of polymer amphiphilicity, and more generally solvation, in CuAAC protocols aimed towards the synthesis of cyclic polymers.

The Covalent Linking of Organophosphorus Heterocycles to Date Palm Wood-Derived Lignin: Hunting for New Materials with Flame-Retardant Potential

Environmentally acceptable and renewably sourced flame retardants are in demand. Recent studies have shown that the incorporation of the biopolymer lignin into a polymer can improve its ability to form a char layer upon heating to a high temperature. Char layer formation is a central component of flame-retardant activity. The covalent modification of lignin is an established technique that is being applied to the development of potential flame retardants. In this study, four novel modified lignins were prepared, and their char-forming abilities were assessed using thermogravimetric analysis. The lignin was obtained from date palm wood using a butanosolv pretreatment. The removal of the majority of the ester groups from this heavily acylated lignin was achieved via alkaline hydrolysis. The subsequent modification of the lignin involved the incorporation of an azide functional group and copper-catalysed azide–alkyne cycloaddition reactions. These reactions enabled novel organophosphorus heterocycles to be linked to the lignin. Our preliminary results suggest that the modified lignins had improved char-forming activity compared to the controls. 31P and HSQC NMR and small-molecule X-ray crystallography were used to analyse the prepared compounds and lignins.

Liquid-crystalline pillar[5]arene-based [2]rotaxanes

ABSTRACT Liquid-crystalline mechanically interlocked molecules have been prepared from a clickable pillar[5]arene-containing [2]rotaxane building block incorporating ethynylated stoppers and dendritic mesogenic subunits of generation 0, 1 and 2 under copper-catalysed alkyne-azide cycloaddition conditions. The liquid-crystalline properties of the resulting pillar[5]arene-based rotaxanes bearing 4, 8 and 16 peripheral cyanobiphenyl units have been investigated and compared to those of corresponding model compounds lacking the macrocyclic component.

Stable light-emitting bionic antibody-conjugated covalent organic frameworks for the adsorption of urea and control of ethyl carbamate

In this paper, a novel azide-modified covalent organic framework conjugated bionic antibody microspheres (COFs(azide)@BAs) were prepared for the adsorption of urea and indirect control of ethyl carbamate (EC) in the processing of Huangjiu. To improve the fluorescence properties and specific binding ability to target, COFs(azides) were synthesized using post-modification of copper-catalyzed azide-alkyne cycloaddition (CuAAC). The aggregation-induced enhancement (AIE) interaction between COFs(azides)@BAs and urea induced an enhanced fluorescence behavior. The detection limit of urea was as low as 13 ng L−1 (R2 = 0.9896) in the linear range of 0.05–5 μg L−1. In addition, a satisfactory control rate for the removal of EC was achieved by COFs(azide)@BAs in the simulated system of Huangjiu processing. Therefore, this research broadened the path for the application of optical sensing systems in food safety control, which had important food safety implication and great economic value.

Experimental and theoretical evaluation of the anticorrosive proprieties of new 1,2,3-triazolyl-acridine derivatives

Three novel 1,2,3-triazolyl-acridine derivatives (abbreviated as TTA, ATM, and ATA) were synthesized via a fast ultrasound-assisted copper-catalyzed azide-alkyne cycloaddition (CuAAC) in good yields. They were studied as corrosion inhibitors for 1020 mild steel in acidic media (1 mol/L HCl) using gravimetric and electrochemical measurements. Weight Loss Study showed that those molecules have anticorrosive efficiency varying from 85 to 94 % at 1 mmol/L (298 K). They also increased their corrosion mitigation at higher temperatures, reaching up to 90–96 % at 338 K. Isotherm fitting revealed that all developed corrosion inhibitors follow the Langmuir theory and data crossing confirmed the monolayer formation. Atomic Force Microscopy suggested the presence of a protective film on the metal surface and Electrochemical Impedance Spectroscopy, Linear Polarization Resistance, and Electrochemical Frequency Modulation showed a better charge transfer and polarization resistance alongside a lower corrosion current density in the presence of TTA, ATM, and ATA in the corrosive media. Also, polarization curves characterized all three organic molecules as mixed-type corrosion inhibitors for mild steel. First-principles density functional theory (DFT) simulations revealed that all molecules form covalent bonds with iron atoms upon adsorption on Fe(110) surface. The ATA molecule exhibited a bond-breaking upon adsorption and had a higher interaction energy with the iron surface. The chemical interactions between inhibitors’ molecules and iron atoms were confirmed by projected density of states analysis, showing a strong hybridization between molecules’ orbitals and 3d iron orbitals.

Transcription-Factor-Induced Aggregation of Biomimetic Oligonucleotide-b-Protein Micelles.

Polymeric micelles and especially those based on natural diblocks are of particular interest due to their advantageous properties in terms of molecular recognition, biocompatibility, and biodegradability. We herein report a facile and straightforward synthesis of thermoresponsive elastin-like polypeptide (ELP) and oligonucleotide (ON) diblock bioconjugates, ON-b-ELP, through copper-catalyzed azide-alkyne cycloaddition. The resulting thermosensitive diblock copolymer self-assembles above its critical micelle temperature (CMT ∼30 °C) to form colloidally stable micelles of ∼50 nm diameter. The ON-b-ELP micelles hybridize with an ON complementary strand and maintain their size and stability. Next, we describe the capacity of these micelles to bind proteins, creating more complex structures using the classic biotin-streptavidin pairing and the specific recognition between a transcription factor protein and the ON strand. In both instances, the micelles are intact, form larger structures, and retain their sensitivity to temperature.